Under “ideal” conditions it is possible to model retention in gradient elution so as to be able to calculate retention times, tG, as a function of isocratic retention in corresponding liquid chromatographic systems. In this paper we consider various “non-ideal” processes that lead to errors in calculated values of tG. 

An HPLC-method for the measurement of blood Cyclosporin A levels (CyA) of renal allograft transplanted patients within 9 min is described. After a simple protein precipitation of the blood the supernatant is transferred to an HPLC-system. The short time of analysis is obtained by a step gradient elution technique and a precolumn separation of the fractions of interest followed by a backflush regeneration step of the precolumn. The analysis of the fraction of interest takes place on a column with high resolution power as long as the precolumn is regenerated. CyA is monitored by UV-absorption at 206 nm. Detection of 20 to 2000 ng/ml Cya allows the use of the method for patient monitoring and for research purposes.

Preliminary model for predicting large-molecule separations by gradient elution, particularly for reversed-phase LC

Reduced plate height (h) vs. reduced velocity (v) plots have been measured over a wide range of v for 36 high-performance liquid chromatographic systems. Column type was varied over wide limits and solute capacity factor (k′) values were changed over the range 0.6–22. Resulting data can be accurately described by the Knox equation h = Av1/3 + B/v + Cv, where A is roughly constant (A = 0.5–0.8) for all columns studied, but values of B and C are strongly dependent on column type and solute k′ values. Reduced plate height (h) vs. reduced velocity (v) plots have been measured over a wide range of v for 36 high-performance liquid chromatographic systems. Resulting data can be accurately described by the Knox equation h = Av1/3 + B/v + Cv, where A is roughly constant (A = 0.5–0.8) for all columns studied, but values of B and C are strongly dependent on column type and solute k′ values. 

Analysis of proteins in solution by high-performance liquid chromatography is presented with respect to structural changes in solution, adsorption processes and differentation concerning specific activities. Trypsin and cellulases were taken as examples.

Development of the basic theory relating gradient and isocratic separations, essential to later work on DryLab I and DryLab G.